The challenge to meet the ever increasing demand for oil includes increasing crude oil recovery from heavy oil reservoirs. This challenge has resulted in expanding efforts to develop alternative cost efficient oil recovery processes (Kianipey, S. A. and Donaldson, E. C. 61st Annual Technical Conference and Exhibition, New Orleans, La., USA, Oct. 5-8, 1986). Heavy hydrocarbons in the form of petroleum deposits and oil reservoirs are distributed worldwide. These oil reservoirs are measured in the hundreds of billions of recoverable barrels. Because heavy crude oil has a relatively high viscosity, it is essentially immobile and cannot be easily recovered by conventional primary and secondary means.
Microbial Enhanced Oil Recovery (MEOR) is a methodology for increasing oil recovery by the action of microorganisms (Brown, L. R., Vadie, A. A., Stephen, O. J. SPE 59306, SPE/DOE Improved Oil Recovery Symposium, Oklahoma, Apr. 3-5, 2000). MEOR research and development is an ongoing effort directed at discovering techniques to use microorganisms to modify crude oil properties to benefit oil recovery (Sunde. E., Beeder, J., Nilsen, R. K. Torsvik, T., SPE 24204, SPE/DOE 8th Symposium on enhanced Oil Recovery, Tulsa, Okla., USA, Apr. 22-24, 1992).
Methods for identifying microorganisms useful in MEOR processes have been described. These methods require identification of samples drawn from an oil well or reservoir comprising a consortium of microorganisms and enrichment or evolution of populations in the sample under specific conditions with a defined nutrient medium (U.S. Patent Application No. 2007/0092930A1). Thus, there is a need for developing methods to: 1) identify microorganisms that can grow in or on oil under anaerobic denitrifying conditions by selection of pure isolates from enrichment of indigenous microorganisms; 2) screen isolates for properties that might be useful in oil modification or interactions and 3) use said identified microorganisms, in a cost-efficient way, to improve oil recovery.
The microorganism described herein has been identified as a strain of Pseudomonas stutzeri. Pseudomonas stutzeri belongs to a broad category of denitrifying bacteria that is found in, and adaptable to, many environments. Pseudomonas stutzeri are grouped into genomovars based on DNA-DNA hybridization. Typing by 16S rDNA gene sequence has been in agreement with the hybridization typing. However, Pseudomonas stutzeri is considered to have high genetic mutation rates (Rius, Nuria, R., et al., J. Bacteriol., 183, 736-744, 2001) and is easily transformed in its natural environment (Sikorski, J., et al., Environ. Microbiol., 4, 456-476, 2002). Pseudomonas stutzeri strain LH4:15 16S rDNA gene has 100% homology to a Pseudomonas stutzeri (strain 24a97) isolated from soil contaminated with mineral oil near a filling station in Northern Germany (Sikorski, J. et al., supra). Other Pseudomonas stutzeri strains have been found in association with oil and petroleum and were seen to degrade alkanes. Aerobic cleavage of C—N bonds in oil compounds by Pseudomonas stutzeri strains has been disclosed (U.S. Pat. No. 6,541,240B1).
Strains of Pseudomonas stutzeri have been used in bioremediation processes. Crude oil and petroleum product bioremediation from water and soil by a consortium that contains Pseudomonas stutzeri is described in WO 95/031408A1. Pseudomonas stutzeri strain JJ anaerobically degrades 2-chloroethanol under denitrifying conditions (Diijk, J. A., et al., Appl. Microbiol. Biotechnol., 63, 68-74, 2003). Strain KC was isolated from an aquifer and transforms carbon tetrachloride to carbon dioxide, formate and other non-volatile compounds anaerobically (Criddle, C. S., et al., Appl. Environ. Microbiol., 56, 3240-3246, 1990). Aerobic biodegradation of aromatic hydrocarbons has been widely observed in Pseudomonas stutzeri strains, but observation of anaerobic degradation has been limited (Lalucat, J., et al., Microbiol. Mol. Biol. Rev. 70: 510-547, 2006). Pseudomonas stutzeri has been used for bioremediation of other xenobiotic, toxic environmental pollutants, e.g., nitrogen compounds, biocides, high molecular weight polyethylene glycols, and metals.
Pseudomonas stutzeri is also known to form biofilms (Viggiani, A., et al., J. Biotechnol. 123, 464-77, 2006). Pseudomonas stutzeri biofilm has been used to remove oxidized selenium from water (U.S. Pat. No. 6,183,644B1), and Pseudomonas stutzeri (accession #MCMRD-AB-001) is disclosed as a biofilm for the production of xylanase (US Patent Application No. 20030008379).